Hematopoietic stem cells (HSC) are multipotent cells that reside in the bone marrow and replenish all adult hematopoietic lineages throughout the lifetime of the animal. While experimenting with staining of murine bone marrow cells with the vital dye, Hoechst 33342, we discovered that display of Hoechst fluorescence simultaneously at two emission wavelengths revealed a small and distinct subset of whole bone marrow cells that had phenotypic markers of multipotential HSC. These cells were shown in competitive repopulation experiments to contain the vast majority of HSC activity from murine bone marrow and to be enriched at least 1,000-fold for in vivo reconstitution activity. Further, these Hoechst-stained side population (SP) cells were shown to protect recipients from lethal irradiation at low cell doses, and to contribute to both lymphoid and myeloid lineages. The formation of the Hoechst SP profile was blocked when staining was performed in the presence of verapamil, indicating that the distinctly low staining pattern of the SP cells is due to a multidrug resistance protein (mdr) or mdr-like mediated efflux of the dye from HSC. The ability to block the Hoechst efflux activity also allowed us to use Hoechst to determine the DNA content of the SP cells. Between 1 and 3% of the HSC were shown to be in S-G2M. This also enabled the purification of the G0-G1 and S-G2M HSC had a reconstitution capacity equivalent to quiescent stem cells. These findings have implications for models of hematopoietic cell development and for the development of genetic therapies for diseases involving hematopoietic cells.

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Isolation and functional properties of murine hematopoietic stem cells that are replicating in vivo.

Background Irradiation after mastectomy can reduce locoregional recurrences in women with breast cancer, but whether it prolongs survival remains controversial. We conducted a randomized trial of radiotherapy after mastectomy in high-risk premenopausal women, all of whom also received adjuvant systemic chemotherapy with cyclophosphamide, methotrexate, and fluorouracil (CMF). Methods A total of 1708 women who had undergone mastectomy for pathological stage II or III breast cancer were randomly assigned to receive eight cycles of CMF plus irradiation of the chest wall and regional lymph nodes (852 women) or nine cycles of CMF alone (856 women). The median length of follow-up was 114 months. The end points were locoregional recurrence, distant metastases, disease-free survival, and overall survival. Results The frequency of locoregional recurrence alone or with distant metastases was 9 percent among the women who received radiotherapy plus CMF and 32 percent among those who received CMF alone (P<0.001). The ...

Postoperative Radiotherapy in High-Risk Premenopausal Women with Breast Cancer Who Receive Adjuvant Chemotherapy

In 1976 we began a randomized trial to evaluate breast conservation by a segmental mastectomy in the treatment of Stage I and II breast tumors less than or equal to 4 cm in size. The operation removes only sufficient tissue to ensure that margins of resected specimens are free of tumor. Women were randomly assigned to total mastectomy, segmental mastectomy alone, or segmental mastectomy followed by breast irradiation. All patients had axillary dissections, and patients with positive nodes received chemotherapy. Life-table estimates based on data from 1843 women indicated that treatment by segmental mastectomy, with or without breast irradiation, resulted in disease-free, distant-disease-free, and overall survival at five years that was no worse than that after total breast removal. In fact, disease-free survival after segmental mastectomy plus radiation was better than disease-free survival after total mastectomy (P = 0.04), and overall survival after segmental mastectomy, with or without radiation, was better than overall survival after total mastectomy (P = 0.07, and 0.06, respectively). A total of 92.3 per cent of women treated with radiation remained free of breast tumor at five years, as compared with 72.1 per cent of those receiving no radiation (P less than 0.001). Among patients with positive nodes 97.9 per cent of women treated with radiation and 63.8 per cent of those receiving no radiation remained tumor-free (P less than 0.001), although both groups received chemotherapy. We conclude that segmental mastectomy, followed by breast irradiation in all patients and adjuvant chemotherapy in women with positive nodes, is appropriate therapy for Stage I and II breast tumors less than or equal to 4 cm, provided that margins of resected specimens are free of tumor.

Five-Year Results of a Randomized Clinical Trial Comparing Total Mastectomy and Segmental Mastectomy with or without Radiation in the Treatment of Breast Cancer

NCCN Clinical Practice Guidelines in Oncology (NCCN Guidelines

Background Radiotherapy after mastectomy to treat early breast cancer has been known since the 1940s to reduce rates of local relapse. However, the routine use of postoperative radiotherapy began t...

Adjuvant Radiotherapy and Chemotherapy in Node-Positive Premenopausal Women with Breast Cancer

In 31 cases of Stages I or II carcinoma of the breast treated by primary radiation therapy, the cosmetic results were analyzed with regard to the details of treatment. Three principal treatment factors were identified which influenced the cosmetic outcome: (1) the extent and location of the biopsy procedure, (2) the time/dose factors of the radiation therapy and (3) the technique of the radiation therapy. Cosmetic results were lessened when the biopsy procedure included a wide resection of adjacent breast tissue or when the biopsy scar was obvious. Increasing doses of external beam radiation were associated with greater degrees of retraction and fibrosis of the treated breast. All 6 patients who received 6000 rad by external beam had significant retraction and fibrosis while patients who received 5000 rad rarely showed significant changes. Local boost doses by interstitial implantation did not diminish the cosmetic outcome. All patients were treated using supervoltage equipment without bolus and skin changes secondary to treatment were infrequent. Seventeen patients developed localized areas of fibrosis and skin changes at the matchline between adjacent radiation fields. Recommendations are made for improved cosmetic results based on these findings.

Analysis of cosmetic results following primary radiation therapy for stages I and II carcinoma of the breast.

Thirty-four gross and histologic features of the primary tumor in 231 cases of clinical Stage I and II invasive breast cancer were reviewed in an attempt to identify features which might correlate with an increased risk of local recurrence within the breast following biopsy and primary radiation therapy. Local recurrence risk at 5 years was calculated for each feature studied. While results are reported in terms of 5-year actuarial local recurrence risk, not all patients were followed for 5 years (median follow-up period, 44 months). Patients with cases in which the biopsy was less than excisional had a considerably greater actuarial risk of local recurrence at 5 years than those in which the biopsy was excisional (36% versus 8%; P = 0.0005). Among 154 infiltrating ductal carcinomas treated by excisional biopsy prior to radiotherapy, histologic features associated with a significantly increased local recurrence risk at 5 years were the combination of extensive intraductal involvement by carcinoma and high nuclear grade and/or high mitotic index. Twenty-seven tumors demonstrated this constellation of features, and there was a 5-year actuarial local recurrence risk of 39% among this group. The local recurrence risk for the remainder of the population was only 4% (P less than 0.0001). Thus, through pathologic examination of the primary, the authors identified a subgroup of patients with a considerably increased risk of local recurrence following biopsy and primary radiotherapy.

https://onlinelibrary.wiley.com/doi/pdf/10.1002/1097-0142%2819840301%2953%3A5%3C1049%3A%3AAID-CNCR2820530506%3E3.0.CO%3B2-O

Pathologic predictors of early local recurrence in Stage I and II breast cancer treated by primary radiation therapy

Between April 1969, and December 1974, 111 consecutive surgically staged I A and II A patients with supradiaphragmatic Hodgkin's disease were treated at the Joint Center for Radiation Therapy. Patients received 3600--4400 rad to mantle and para-aortic--splenic pedicle regions. Median follow-up was 56 months (30--96). Fourteen patients developed relapsing Hodgkin's disease and three patients died of possible treatment-related causes, two with acute myocardial infarctions and one with radiation pneumonitis. Patients with mediastinal enlargement greater than one third of the chest diameter have a significantly higher risk (p less than 0.01) of developing relapse (9 of 18) than patients with lesser or no mediastinal disease (5 of 93). Of the 18 patients with large mediastinal disease, six relapsed in the mediastinum and two in the lung. There continue to be no pelvic extensions in the entire group. There is a 92% relapse-free and 97% overall survival in the 93 patients without extensive mediastinal disease. We continue to recommend mantle and para-aortic--splenic pedicle irradiation for these patients. In view of the large number of relapses in patients with extensive mediastinal disease, we are now treating this subgroup of patients with MOPP chemotherapy in addition to mantle and para-aortic irradiation.

https://acsjournals.onlinelibrary.wiley.com/doi/pdf/10.1002/1097-0142%28197809%2942%3A3%3C1039%3A%3AAID-CNCR2820420302%3E3.0.CO%3B2-R

The significance of mediastinal involvement in early stage Hodgkin's disease.

Primary radiation therapy has become an accepted alternative to mastectomy for patients with early breast cancer. In order to improve the results of this treatment, we performed an analysis of failure on 366 clinical Stage I or II invasive breast carcinomas treated with primary radiation therapy. With a median follow-up of 52 months, there have been 30 recurrences in the treated breast, for a 5 year actuarial local recurrence rate of 9%. The recurrence rate was much higher for patients having less-than-excisional biopsy than for those undergoing excisional biopsy (35% vs. 7%, p < 0.0001). Failures in the treated breast have been categorized as: true recurrence (TR), directly at the site of the primary; marginal miss (MM), at the edge of the boosted volume; and elsewhere in the breast (E). For patients having excisional biopsy, the actuarial probability of a true recurrence at 5 years was 4%. The risk of a true recurrence was related to the dose given to the primary site with rare true recurrences seen with doses greater than 6000 rad. The probability of a marginal miss at 5 years was 4%, and was not related to the dose or volume of the boost used. The probability of a recurrence elsewhere in the breast at 5 years was 1%. In patients treated with excisional biopsy and a total primary dose over 6000 rad, the probability of a local recurrence at 5 years was 4 and 10% for clinical Stage I and II patients respectively. These results indicate that primary radiation therapy provides a high likelihood of local tumor control and our analysis of failure suggests methods for decreasing each type of local recurrence.

Breast relapse following primary radiation therapy for early breast cancer. I. Classification, frequency and salvage.

In order to assess the cosmetic results of treatment, the results in 239 patients with early breast cancer treated by primary radiation treatment without adjuvant chemotherapy were reviewed. Four patients had bilateral cancers, making a total of 243 breasts available for analysis. Follow-up ranged from 24 to 78 months with a median of 33 months. The parameters measured were breast edema, retraction, telangiectasia, arm edema, and the overall cosmetic appearance. The cosmetic results declined over the first 3 years after treatment, but then stabilized. At 5 years, the overall cosmetic results were judged by physicians as excellent in 77%, good in 9%, fair in 9%, and poor in 5%. A fair or poor cosmetic result was highly correlated with the development of moderate or severe breast retraction. Telangiectasia was uncommonly the only cause of a fair or poor cosmetic result. Breast and arm edema were rarely noted to be significant, but were more common in patients who underwent axillary dissection. In 210 cases, a supplementary boost dose of radiation was delivered to the primary tumor area, and in 33 cases a boost was not used. This boost consisted of an interstitital iridium-192 implant in 204 cases and either high-energy photons or electrons in the remainder. At 4 years, no patient treated without a boost had a fair or poor result compared with 22% who received a boost (P = 0.13). The conclusion is that, in general, primary radiation treatment provides highly satisfactory cosmetic results for patients with early breast cancer.

Samuel Hellman

Papers

Analysis of cosmetic results following primary radiation therapy for stages I and II carcinoma of the breast.

Pathologic predictors of early local recurrence in Stage I and II breast cancer treated by primary radiation therapy

The significance of mediastinal involvement in early stage Hodgkin's disease.

Breast relapse following primary radiation therapy for early breast cancer. I. Classification, frequency and salvage.

Cosmetic results following primary radiation therapy for early breast cancer